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Riding the spermatogenic wave: profiling gene expression within neonatal germ and sertoli cells during a synchronized initial wave of spermatogenesis in mice.

ABSTRACT: Continual sperm production relies on germ cells undergoing spermatogenesis asynchronously. As a result, the testis always contains a mixed population of germ cells at different stages of their differentiation process. The heterogeneous nature of the testis makes profiling gene expression within Sertoli cells or specific populations of germ cells impossible when a wild-type testis is assessed. We recently reported a unique method for synchronizing spermatogenesis without affecting fertility by manipulating RA levels within the neonatal testis. Using this protocol, combined with the RiboTag transgenic mouse line, we have mapped the Sertoli and germ cell translatome during the initial synchronized wave of spermatogenesis. Using microarray analysis, we identified 392 and 194 germ cell and Sertoli cells transcripts, respectively, that dynamically change during spermatogonial differentiation, division, and the onset of meiosis. Functional annotation clustering revealed that transcripts enriched in germ cells were mostly associated with meiosis (21 transcripts), chromatin organization (12 transcripts), and cell cycle (3 transcripts). In addition, glycoproteins (65 transcripts), cell adhesion (15 transcripts), and cell junction (13 transcripts) transcripts were overrepresented in the Sertoli cell-enriched list. These datasets represent the first transcriptional analysis of spermatogonial differentiation, division, and meiotic onset. These data suggest that several of the genes encoding meiotic proteins are expressed and are actively being translated well before germ cells enter meiosis. In addition, this study provides novel candidate genes, Asf1b and Esyt3, that may be involved in the regulation of spermatogonial chromatin reorganization, germ-Sertoli cell interactions, and/or blood-testis barrier formation.

SUBMITTER: Evans E

PROVIDER: S-EPMC4076369 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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Riding the spermatogenic wave: profiling gene expression within neonatal germ and sertoli cells during a synchronized initial wave of spermatogenesis in mice.

Evans Elizabeth E Hogarth Cathryn C Mitchell Debra D Griswold Michael M

Biology of reproduction 20140409 5

Continual sperm production relies on germ cells undergoing spermatogenesis asynchronously. As a result, the testis always contains a mixed population of germ cells at different stages of their differentiation process. The heterogeneous nature of the testis makes profiling gene expression within Sertoli cells or specific populations of germ cells impossible when a wild-type testis is assessed. We recently reported a unique method for synchronizing spermatogenesis without affecting fertility by ma ...[more]

PMID: 24719255

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Project description:BackgroundInfertile men with non-obstructive azoospermia (NOA) have impaired spermatogenesis. Dilated and un-dilated atrophic seminiferous tubules are often present in the testes of these patients, with the highest likelihood of active spermatogenesis in the dilated tubules. Little is known about the un-dilated tubules, which in NOA patients constitute the majority. To advance therapeutic strategies for men with NOA who fail surgical sperm retrieval we aimed to characterize the spermatogonial stem cell microenvironment in atrophic un-dilated tubules.MethodsTestis biopsies approximately 3x3x3 mm3 were obtained from un-dilated areas from 34 patients. They were classified as hypospermatogenesis (HS) (n=5), maturation arrest (MA) (n=14), and Sertoli cell only (SCO) (n= 15). Testis samples from five fertile men were included as controls. Biopsies were used for histological analysis, RT-PCR analysis and immunofluorescence of germ and Sertoli cell markers.ResultsAnti-Müllerian hormone mRNA and protein expression was increased in un-dilated tubules in all three NOA subtypes, compared to the control, showing an immature state of Sertoli cells (p<0.05). The GDNF mRNA expression was significantly increased in MA (P=0.0003). The BMP4 mRNA expression showed a significant increase in HS, MA, and SCO (P=0.02, P=0.0005, P=0.02, respectively). The thickness of the tubule wall was increased 2.2-fold in the SCO-NOA compared to the control (p<0.05). In germ cells, we found the DEAD-box helicase 4 (DDX4) and melanoma-associated antigen A4 (MAGE-A4) mRNA and protein expression reduced in NOA (MAGE-A: 46% decrease in HS, 53% decrease in MA, absent in SCO). In HS-NOA, the number of androgen receptor positive Sertoli cells was reduced 30% with a similar pattern in mRNA expression. The γH2AX expression was increased in SCO as compared to HS and MA. However, none of these differences reached statistical significance probably due to low number of samples.ConclusionsSertoli cells were shown to be immature in un-dilated tubules of three NOA subtypes. The increased DNA damage in Sertoli cells and thicker tubule wall in SCO suggested a different mechanism for the absence of spermatogenesis from SCO to HS and MA. These results expand insight into the differences in un-dilated tubules from the different types of NOA patients.

Dataset Information

Riding the spermatogenic wave: profiling gene expression within neonatal germ and sertoli cells during a synchronized initial wave of spermatogenesis in mice.

Publications

Riding the spermatogenic wave: profiling gene expression within neonatal germ and sertoli cells during a synchronized initial wave of spermatogenesis in mice.

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